The long-term goals of this project are to isolate, characterize and study the expression of the human actin genes in normal and pathological muscle and cell development. This will allow us to approach an understanding of how the human organism utilizes the actin multi-gene family to produce the various saromeric and cytoskeletal structures which result from actin gene expression. We have already learned that there are more than 25 human actin genes. These genes will be isolated by recombinant DNA technology from banks of genomic and cDNA clones. We will obtain a detailed picture of the topology of the actin genes with respect to non-coding region and promoter homologies and the presence of intervening sequences and repetitive sequences. We will be able to relate these to the type of actin encoded by these genes. This will also provide valuable information concering the number of alpha-, beta- and gamma-actin genes and their evolution. In addition, we will obtain probes with which to map their chromosome location and information to enable interpretation of actin gene polymorphism in the human population. We will investigate the possible use of actin polymorphism to diagnose genetic diseases. The isolation and characterization of the human actin genes will provide a knowledge base which should allow us to identify at a molecular level the cause of any actin gene lesions. The number of different genes which are utilized by human cells to produce the cytoplasmic actins will be determined. Comparisons between different cell types will allow us to determine whether the expression of genes encoding the cytoplasmi actins under any tissue specific regulation. The types of actin proteins expresses in non-muscle cells will be determined. In collaboration with the laboratory of Dr. Helen Blau, we will measulre the relative levels of alpha-, beta- and gamma-actin mRNA transcripts and the kinetics of their synthesis during human myogenesis in cell culture. We will also examine the genomic organization of the actin genes during myogenesis in order to evaluate possible changes in actin gene structure and number. This will lprovide us with a sytem in which to study the mechanisms by which human muscle induces the synthesis of alpha-actin and ceases synthesis of beta- and gamma-actin.